Salmonella is a rod-shaped, Gram-negative Enterobacteria. Salmonella are closely related to the Escherichia genus and are found worldwide in warm- and cold-blooded animals, in humans. They cause illnesses in humans and many animals, such as typhoid fever, paratyphoid fever, and the food-borne illness, salmonellosis. Most persons infected with Salmonella develop diarrhea, fever, vomiting, and abdominal cramps 12 to 72 hours after infection. Infection is usually diagnosed by culture of a stool sample. The illness usually lasts 4 to 7 days. Although most people recover without treatment, severe infections can occur. Infants, elderly people, and those with weakened immune systems are more likely than others to develop severe illness. When severe infection occurs, Salmonella may spread from the intestines to the bloodstream and then to other body sites and can cause death unless the person is treated promptly with antibiotics.
Currently, there are two recognized Salmonella species: S. enterica and S. bongori, with six main serotypes: enterica (I), salamae (II), arizonae (IIIa), diarizonae (IIIb), houtenae (IV), and indica (VI). The presence of several pathogenicity islands (PAIS) that encode various virulence factors allows Salmonella spp. to colonize and infect host organisms. There are two important PAIs, Salmonella pathogenicity island 1 and 2 (SPI-1 and SPI-2) that encode two different type III secretion systems for the delivery of effector molecules into the host cell that result in internalization of the bacteria which then leads to systemic spread. Salmonella bongori is reptile-specific and is rarely found in human infections. Certain serovars of Salmonella enterica are responsible for more serious diseases such as Typhoid fever. Salmonella contamination of various foods and surfaces is recognized as a major health hazard by the United States Department of Agriculture Food Safety Inspection Service (FSIS). Salmonella is not destroyed by freezing.
Traditionally, Salmonella in food and on surfaces was detected by performing classical microbiological assays which are time consuming, labor intensive, insensitive, expensive and difficult to automate. With the advent of DNA amplification technologies, Salmonella is now routinely detected by nucleic acid analysis of raw and processed food samples or surface wipe tests. Nucleic acid assays are now adopted by most government agencies as the method of choice to monitor food processing facilities and distribution centers. Nucleic acid analysis permits not only the detection of Salmonella, the identification of the serotype but also the tracking of a Salmonella outbreak. Such monitoring improves public health and safety by putting hospitals and the medical community on notice and also allows for the rapid identification and isolation of the source of the contamination.
For example, in 2009, the FDA announced they had traced the source of an outbreak of Salmonella typhimurium to a plant in Blakely, Ga., owned by Peanut Corporation of America (PCA), and urged people to postpone eating commercially-prepared or manufactured peanut butter-containing products and institutionally-served peanut butter. Salmonella was reported to be found in 46 states in the United States in at least 3,862 peanut butter-based products such as crackers, energy bars, and peanut butter cookies from at least 343 food companies. Dog treats were affected as well. At least 691 people in more than 46 states became sick, and the Salmonella claimed at least nine lives. Peanut butter and peanut paste manufactured by PCA were distributed to hundreds of firms for use as an ingredient in thousands of different products, such as cookies, crackers, cereal, candy and ice cream, all of which were recalled. Some products were also sold directly to consumers in retail outlets. Containment of a Salmonella outbreak therefore presents governmental agencies with a daunting, logistical task.
A prompt and comprehensive response to a Salmonella outbreak undoubtedly saves money, widespread illness and even lives. Nevertheless, PCR amplification methods are prone to the detection of false-positives (as described in WO 95/33854) and false-negatives (as described in WO 92/01056, WO 95/00664, WO 92/01056, WO 93/04202). For the foregoing reasons, there is still a unmet need in the art for the accurate and reliable real-time detection of Salmonella nucleic acid sequences in test samples.